US7302913B2 - Vaccine against salmonid rickettsial septicaemia based on arthrobacter cells - Google Patents

Vaccine against salmonid rickettsial septicaemia based on arthrobacter cells Download PDF

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US7302913B2
US7302913B2 US10/521,104 US52110405A US7302913B2 US 7302913 B2 US7302913 B2 US 7302913B2 US 52110405 A US52110405 A US 52110405A US 7302913 B2 US7302913 B2 US 7302913B2
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arthrobacter
fish
salmonis
vaccine
renogen
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US20050129714A1 (en
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Kira Salonius
Steven Gareth Griffiths
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Elanco Tiergesundheit Ag F/k/a Novartis Tiergesundheit Ag
Elanco Tiergesundheit AG
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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K39/00—Medicinal preparations containing antigens or antibodies
    • A61K39/02—Bacterial antigens
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K39/00—Medicinal preparations containing antigens or antibodies
    • A61K39/02—Bacterial antigens
    • A61K39/0233—Rickettsiales, e.g. Anaplasma
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K39/00—Medicinal preparations containing antigens or antibodies
    • A61K39/02—Bacterial antigens
    • A61K39/05—Actinobacteria, e.g. Actinomyces, Streptomyces, Nocardia, Bifidobacterium, Gardnerella, Corynebacterium; Propionibacterium
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P31/00—Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
    • A61P31/04—Antibacterial agents
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K39/00—Medicinal preparations containing antigens or antibodies
    • A61K2039/51—Medicinal preparations containing antigens or antibodies comprising whole cells, viruses or DNA/RNA
    • A61K2039/52—Bacterial cells; Fungal cells; Protozoal cells
    • A61K2039/521—Bacterial cells; Fungal cells; Protozoal cells inactivated (killed)
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K39/00—Medicinal preparations containing antigens or antibodies
    • A61K2039/555—Medicinal preparations containing antigens or antibodies characterised by a specific combination antigen/adjuvant
    • A61K2039/55588—Adjuvants of undefined constitution
    • A61K2039/55594—Adjuvants of undefined constitution from bacteria
    • Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
    • Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
    • Y02A—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE
    • Y02A50/00—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE in human health protection, e.g. against extreme weather
    • Y02A50/30—Against vector-borne diseases, e.g. mosquito-borne, fly-borne, tick-borne or waterborne diseases whose impact is exacerbated by climate change

Definitions

  • the present invention concerns use of a live strain of Arthrobacter in the preparation of a medicament to treat or prevent salmonid rickettsial septicaemia (SRS), and vaccines based on these bacteria.
  • SRS salmonid rickettsial septicaemia
  • Piscirickettsia salmonis is a gram-negative obligate intracellular bacterium that causes systemic septicaemia (salmonid rickettsial syndrome, SRS, or piscirickettsiosis) in salmonid fish.
  • Piscirickettsia -like bacteria are now been recognized with increasing frequency in a variety of other fish species, from both fresh and salt waters around the world.
  • Piscirickettsiosis and piscirickettsiosis-like diseases have affected aquaculture productivity, profitability, the species compatible with commercial rearing, and transportation of fish from site to site.
  • the Chilean aquaculture industry alone attributes annual losses to salmonid piscirickettsiosis of $150 million. In Chile, the syndrome has led to a shift from the more commercially desirable coho salmon to the less desirable but more piscirickettsiosis resistant Atlantic salmon as the primary cultivated species.
  • Antimicrobials have been tested as a therapy for SRS, but without consistent success. Other suggested measures include attempts to reduce stress in the fish by reducing stocking density, and removing dead fish from tanks without delay. The most practical solution to the SRS epidemic would be to find an effective vaccine to prevent the disease in the first place. Inactivated bacterin preparations from P. salmonis have been shown to have some protective effect, and may be the only suitable option for co-administration in multivalent oil preparations, but are relatively expensive to produce on a commercial scale. Vaccines based on recombinant antigens from P. salmonis have not yet reached the marketplace.
  • the present invention is based on the surprising discovery that an existing commercial vaccine product is remarkably effective in preventing the disease.
  • This vaccine is marketed under the name RENOGEN, a live, non-virulent strain of Arthrobacter vaccine.
  • RENOGEN a live, non-virulent strain of Arthrobacter vaccine.
  • This vaccine is indicated to protect salmon and other farmed fish against bacterial kidney disease (BKD).
  • BKD bacterial kidney disease
  • a medicament for the treatment or prevention of piscirickettsiosis in fish.
  • the preferred targets of the medicament are salmonid fish exposed to risk of SRS infection.
  • the Arthrobacter cells are preferably from the strain deposited under accession number ATCC 55921, or an equivalent strain.
  • a vaccine composition comprising live Arthrobacter cells and a killed bacterial immunostimulant, and a pharmaceutically acceptable carrier.
  • a vaccine composition comprising killed Arthrobacter cell material, and use of killed Arthrobacter cell material as an immunostimulant.
  • the killed Arthrobacter cell material is preferably from the strain deposited under accession number ATCC 55921, or an equivalent strain.
  • a vaccine composition comprising live Arthrobacter cells and inactivated Piscirickettsia salmonis antigen, whereby the vaccine is optionally provided in the form of a kit comprising a lyophilized Arthrobacter live cell culture and a sterile diluent comprising the inactivated P. salmonis antigen.
  • a method of treatment or prevention of piscirickettsiosis in fish comprising administering to fish in need of such treatment a vaccine comprising live Arthrobacter cells.
  • RENOGEN vaccine has been in use for some time to combat Bacterial Kidney Disease (BKD) in salmonid fish.
  • BKD Bacterial Kidney Disease
  • This vaccine is unique in that it is the first live culture to have been licensed for use in aquaculture, and comprises a live culture of Arthrobacter sp. nov., deposited under Accession No ATCC 55921 with the American Type Culture Collection (10801 University Boulevard, Manassas, Va. 20110-2209) on 20 Dec. 1996.
  • Arthrobacter is not pathogenic to fish; nor is it the causative agent of BKD (which is Renibacterium salmoninarum ).
  • test groups that had received the RENOGEN vaccine exhibited extremely low mortality rates after 471 dd (degree days), amounting to between 88 and 100 relative percent survival (RPS). Even after 1441 dd (equivalent to one year in sea water) the test groups had a RPS of between 69 and 85%, compared to only 48.6% in the inactivated P. salmonis “gold standard” group.
  • RENOGEN is more effective than any other known vaccine in preventing SRS.
  • Live Arthrobacter bacteria are known to be able to enter cells and replicate for a limited period of time. The present inventors believe that this permits the antigen processing of both carbohydrate and protein antigens with sufficient homology to T-cell epitopes of P. salmonis to provide a high level of protection to direct challenge with virulent P. salmonis.
  • the invention therefore provides for the use of Arthrobacter cells in the preparation of a medicament for the treatment or prevention of piscirickettsiosis in fish, in particular salmonid fish, including salmon and trout species, particularly Coho salmon ( Oncorhyncus kisutch ), Chinook salmon ( Oncorhynchus tshawytscha ), masu salmon ( Oncorhyncus masou ), pink salmon ( Oncorhynchus gorbuscha ), rainbow trout ( Oncorhynchus mykiss ), and Atlantic salmon ( Salmo salar ).
  • Coho salmon Oncorhyncus kisutch
  • Chinook salmon Oncorhynchus tshawytscha
  • masu salmon Oncorhyncus masou
  • pink salmon Oncorhynchus gorbuscha
  • rainbow trout Oncorhynchus mykiss
  • Atlantic salmon Salmo salar
  • any other fish species susceptible to piscirickettsiosis or similar disease may benefit, such as, Tilapia sp., Black seabass ( Dicentrarchus sp.), White seabass ( Atractoscion nobilis ), grouper fish, cichlids etc.
  • RenogenTM is based on a particular deposited strain of Arthrobacter (ATCC 55921). In performing the present invention, this strain or equivalent Arthrobacter strains can be employed. Equivalent Arthrobacter strains share the identifying characteristics of Arthrobacter ATCC 55921. They display similar protective capabilities against SRS. A species of Arthrobacter having an identical 16S rDNA sequence or a 16S rDNA sequence having a divergence of less than 3% with the strain ATCC 55921 is regarded as being equivalent. This 16S rDNA sequence is deposited under Genbank accession number AF099202. Another method of defining an equivalent strain is by RAPD assay using the F12-373 primer (5′-ACGGTACCAG-3′), as described in Griffiths, S G et al.
  • SRS immunogenic properties can be identified by the screening assays described in the preceding paragraph and/or by the experimental procedures described in Example 1 and Example 2.
  • the preferred route of administration of the vaccine is by injection into the peritoneal cavity but other administration options exist, including orally in feed, by intra-dermal or intra-muscular injection, or by immersion in sea water or in fresh water. Fish are usually anaesthetized before receiving the vaccine by injection. It is recommended that fish be 10 grams or greater in body weight for administration of the vaccine of the invention by intraperitoneal injection. For immersion or oral administration, a body weight of at least 2 grams is preferred.
  • the effective dosage of vaccine may vary depending on the size and species of the subject, and according to the mode of administration.
  • the optimal dosage can be determined through trial and error by a veterinarian or aquaculture specialist.
  • a suitable dosage range may be from about 10 2 to 10 9 cfu per unit dose, preferably about 10 4 to 10 7 cfu per unit dose, more preferably about 10 5 to 10 6 cfu per unit dose, and most preferably about 10 5 cfu per unit dose. However, higher or lower doses may also be effective.
  • Preferably a single dosage unit is administered to the fish to be treated. Smaller fish may benefit from a dose of about 10 4 to 10 7 cfu/ml with dip (immersion) administration, for instance with a contact time of about 60 seconds.
  • the vaccine may be diluted in 1 to 10 volumes of water before adding to tanks or cages holding fish.
  • a preferred dosage volume for injections is about 0.05-0.5 ml, preferably 0.075-0.25 ml, more preferably 0.1-0.2 ml, optionally about 0.1 ml.
  • live Arthrobacter cells are combined with a pharmaceutically acceptable carrier or vehicle in a pharmaceutical composition.
  • suitable carriers/vehicles include conventional excipients, and may be, for example, solvents such as water, oil, or saline, dextrose, glycerol, wetting or emulsifying agents, bulking agents, coatings, binders, fillers, disintegrants, diluents, lubricants, pH buffering agents, or conventional adjuvants such as muramyl dipeptides, pyridine, aluminium hydroxide, oils, (e.g. mineral oil), saponins, block co-polymers and other substances known in the art.
  • a preferred pharmaceutical composition comprises a saline diluent.
  • vaccines are prepared as liquid solutions or suspensions for injection or for delivery in water.
  • Solid (e.g. powder) forms suitable for dissolution in, or suspension in, liquid vehicles, or for mixing with solid food prior to administration may also be prepared.
  • the vaccine is a lyophilised culture.
  • the vaccine is suitable for reconstitution with a sterile diluent.
  • lyophilized cells may be reconstituted in 0.9% saline (optionally provided as part of the packaged vaccine product).
  • the pharmaceutical vaccine compositions of the invention may be administered in a form for immediate release or extended release.
  • the Arthrobacter vaccine of the invention comprises an immunostimulant.
  • the immunostimulant may be any known immunostimulant, but it is preferably a killed bacterial preparation.
  • the immunostimulant is killed Arthrobacter cell material, which is optionally heat killed and is optionally from a culture of Arthrobacter ATCC 55921.
  • killed bacterial preparations include: “Peptimune” (a heat-killed Arthrobacter ATCC 59921 culture) and “Ultracorn” (ultrasonicated Corynebacterium cutis lysate).
  • An optimal dosage of killed bacterial immunostimulant is (per vaccine unit dose) 1 to 100 ⁇ g, preferably in the range 5 to 50 ⁇ g, more preferably 10 to 20 ⁇ g and optionally about 12 ⁇ g of cellular mailer.
  • the killed bacterial immunostimulant is optionally dissolved or suspended in sterile diluent (e.g. saline) for mixing with lyophilized live Arthrobacter cells.
  • the invention in one aspect provides a vaccine composition comprising live Arthrobacter cells and further comprising at least one other immunogen (where an “immunogen” is defined as a molecule such as an antigen capable of raising a specific immune response in a fish).
  • the immunogen is optionally selected from the group consisting of: inactivated antigen prepared from Piscirickettsia salmonis ( P. salmonis ); a recombinant P. salmonis antigen; and a nucleic acid vector carrying an expressible P. salmonis antigen.
  • RENOGEN vaccine of the invention with a conventional SRS vaccine ( P.salmonis bacterin or recombinant antigen vaccine or nucleic acid vaccine) in a kit comprising both components for separate, sequential or simultaneous administration, for treatment or prevention of SRS.
  • SRS vaccine P.salmonis bacterin or recombinant antigen vaccine or nucleic acid vaccine
  • the invention relates to a vaccine comprising live Arthrobacter cells and inactivated P. salmonis antigen, and optionally killed Arthrobacter cell material as an immunostimulant.
  • the P. salmonis antigen can be prepared by inactivation using any known inactivating agent, but is preferably prepared by formalin inactivation.
  • P. salmonis antigen can be prepared from any isolate of the bacteria.
  • strain LF-89 deposited under ATCC number VR-1361, or a strain derived therefrom, is used to prepare the inactivated antigen.
  • a suitable procedure for inactivating the P. salmonis antigen is by harvesting the supernatant from P. salmonis infected CHSE-14 cell cultures and adding formalin (37% formaldehyde solution) to a final concentration of 0.125% (v/v).
  • the culture fluid/formalin mixture is stirred to homogeneity and then held at 4 ⁇ 2° C. with constant agitation for a minimum of 72 hours.
  • the inactivated harvest material may be concentrated by sterile ultra-filtration.
  • a suitable final concentration of the P. salmonis antigen preparation defined by an enzyme immunoassay (EIA) ratio expressed as OD 405/490 of the antigen/OD 405/490 standard, is 1.5 ⁇ 0.2 units.
  • this vaccine is produced and sold in the form of a kit comprising a lyophilized culture of live Arthrobacter cells, together with a sterile diluent such as saline in which the inactivated P. salmonis antigen (and optionally a killed bacterial immunostimulant) is dissolved or suspended.
  • a sterile diluent such as saline in which the inactivated P. salmonis antigen (and optionally a killed bacterial immunostimulant) is dissolved or suspended.
  • the P. salmonis antigen prepared made as described above can be mixed with the diluent at a concentration of between about 10 to about 150 ml/litre, preferably about 20 to about 100 ml/litre, and most preferably 75 ml/litre.
  • multivalent vaccines comprising live Arthrobacter cells and antigens from pathogens other than P. salmonis.
  • the blot was incubated with 20 ⁇ l of rabbit anti- Arthrobacter polyclonal antibodies for 45 minutes in 15 ml of 1% casein tris-borate saline (cTBS). The blot was then exposed to 5 ⁇ l of goat anti-rabbit immunoglobulin alkaline phosphatase (GAR-AP), and developed. Several proteins were highlighted on the blot, indicating that anti- Arthrobacter protein antibodies have a strong affinity to certain P. salmonis proteins. This result was also confirmed on a 2D Western blot.
  • cTBS casein tris-borate saline
  • the bacterin was prepared from the supernatant of a P. salmonis type strain LF-89 infected CHSE-14 cell culture using 0.125% formalin at 4° C. over a minimum 72 h period. U/F concentration was employed and the concentrated supernatant was used to incorporate 8 ⁇ g (protein) per 0.1 ml dose.
  • the bacterin vaccine was delivered with ULTRACORN (Virbac, France) at 20 (Group 6) and 100 ⁇ g (Group 7) per fish.
  • the antigens were emulsified with an equal volume of mineral oil adjuvant prior to injection.
  • the negative control group (Group 8) received an injection of saline.
  • Table 1 summarizes the treatment groups (dose volume (0.1 mL per fish) for 20 mls):
  • TCID 50 was determined on 96 well plates containing confluent CHSE-214 cells.
  • Table 2 Mortality during the 28 d safety test, maintained at 9-12° C. through-out the safety and pre-challenge period.
  • Table 3 Cumulative Mortality and Relative Percent Survival of Coho salmon (mean weight 10 g) 471 dd post-vaccination with Arthrobacter sp. nov cells (Groups 1-5), Inactivated SRS vaccines, or saline when challenged with virulent P. salmonis by intraperitoneal injection (TCID 50 3 ⁇ 102.9 per fish) at 12° C.
  • the TCID 50 of the fish sampled from the RENOGEN group was lower than the inactivated vaccine group, and both were lower than the saline controls. This is not of apparent clinical relevance, as the contribution of the high titre groups negates the lower infective dosages when averaging.
  • the RENOGEN group did have the lowest percent positives ( ⁇ 20%) as samples with less than 10 2 were considered not to be clinically infected with SRS. This compares to the same samples from the saline control group where 50% of the fish were positive for SRS, and favourably to the inactivated vaccine group with 44% of the fish positive for SRS.
  • Table 5 Cumulative Mortality and Relative Percent Survival of Coho salmon (mean weight 10 g) 1441 dd post-vaccination with Arthrobacter sp. nov cells (Groups 1-5), Inactivated SRS vaccines, or saline when challenged with virulent P. salmonis by intraperitoneal injection (TCID 3 ⁇ 102.9 per fish) at 12° C.
  • RENOGEN provided significant protection against direct challenge with P. salmonis at 471 dd and at 1441 dd post-vaccination.
  • the vaccine was superior to the protection provided by the standard oil vaccine.
  • the study demonstrates that Arthrobacter sp. nov. live vaccine provides a high degree of protection against P. salmonis infection, and that the protective effect is shown to be long-term. Inclusion of a killed Arthrobacter preparation in the vaccine had an immune-stimulating effect resulting in improved survival rates.

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US10/521,104 2002-07-15 2005-02-14 Vaccine against salmonid rickettsial septicaemia based on arthrobacter cells Active 2025-04-27 US7302913B2 (en)

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GBGB0216414.3A GB0216414D0 (en) 2002-07-15 2002-07-15 Organic compounds
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GB0220100A GB0220100D0 (en) 2002-08-29 2002-08-29 Organic compounds
GB02201002 2002-08-29
PCT/EP2003/007605 WO2004006953A2 (fr) 2002-07-15 2003-07-14 Vaccin contre la septicemie rickettsienne des salmonides

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US20090162391A1 (en) * 2007-12-19 2009-06-25 Schering-Plough Animal Health Corporation Vaccine antigens from piscirickettsia salmonis
US20100330113A1 (en) * 2007-12-19 2010-12-30 Intervet International B.V. Vaccine Antigens
US9492521B2 (en) 2014-05-28 2016-11-15 Auburn University Vaccines for control of epidemic Aeromonas hydrophila generated by markerless gene deletion
US20210368745A1 (en) * 2017-12-21 2021-12-02 Gis Gas Infusion Systems Inc. Method and apparatus for fish farms

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WO2013084169A2 (fr) * 2011-12-07 2013-06-13 Yanez Carcamo Alejandro Javier Nouveau substrat de bouillon et substrats solides exempts de sang pour la culture du pathogène de poisson piscirickettsia salmonis
CN105039221B (zh) * 2015-08-07 2019-04-16 中国农业科学院哈尔滨兽医研究所 一株冷水鱼益生菌节杆菌菌株及其用途
NO20200827A1 (en) * 2017-12-26 2020-07-14 Univ Chile Vaccine formulation for fish based on lipid nanovesicles,particularly a proteoliposome or cochleate, with activity against the salmonid rickettsial syndrome (srs)
KR102200721B1 (ko) * 2019-11-18 2021-01-13 (주)애드바이오텍 연어 리케차성 패혈증 예방 또는 치료용 난황항체의 제조방법

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US20030165526A1 (en) * 1999-09-17 2003-09-04 Kuzyk Michael A. Vaccines and agents for inducing immunity against rickettsial diseases, and associated preventative therapy
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US20090162391A1 (en) * 2007-12-19 2009-06-25 Schering-Plough Animal Health Corporation Vaccine antigens from piscirickettsia salmonis
US7811583B2 (en) 2007-12-19 2010-10-12 Intervet International B.V. Antigens and vaccines against Piscirickettsia salmonis
US20100330113A1 (en) * 2007-12-19 2010-12-30 Intervet International B.V. Vaccine Antigens
US20110070256A1 (en) * 2007-12-19 2011-03-24 Intervet International B.V. Vaccine Antigens from Piscirickettsia Salmonis
US9492521B2 (en) 2014-05-28 2016-11-15 Auburn University Vaccines for control of epidemic Aeromonas hydrophila generated by markerless gene deletion
US20210368745A1 (en) * 2017-12-21 2021-12-02 Gis Gas Infusion Systems Inc. Method and apparatus for fish farms

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WO2004006953A3 (fr) 2004-02-19
JP4381977B2 (ja) 2009-12-09
US7707970B2 (en) 2010-05-04
DK1523330T3 (da) 2009-11-23
HK1079100A1 (en) 2006-03-31
WO2004006953A2 (fr) 2004-01-22
JP2005536500A (ja) 2005-12-02
ES2330738T3 (es) 2009-12-15
HK1075840A1 (en) 2005-12-30
BR0312632B1 (pt) 2014-09-16
NO338944B1 (no) 2016-10-31
AU2003250944B2 (en) 2006-11-23
CN1292795C (zh) 2007-01-03
PT1523330E (pt) 2009-09-29
ATE442162T1 (de) 2009-09-15
EP1523330B1 (fr) 2009-09-09
AU2003250944A1 (en) 2004-02-02
CN1668328A (zh) 2005-09-14
US20090155313A1 (en) 2009-06-18
EP1523330A2 (fr) 2005-04-20
CA2491524C (fr) 2012-09-25
NO20050778L (no) 2005-02-14
BR0312632A (pt) 2005-04-19
CA2491524A1 (fr) 2004-01-22
SI1523330T1 (sl) 2010-01-29
US20050129714A1 (en) 2005-06-16

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